Peterson, J. A. | Search bioRxiv

bioRxiv2025

Potent broad-spectrum anti-coronaviral frameshift inhibitors from virtual screen of RNA binding

Coronavirus genomes contain an RNA pseudoknot that directs -1 programmed ribosomal frame-shifting (-1 PRF) to control expression of viral proteins crucial for replication. Ligands that inhibit -1 PRF can thus attenuate viral propagation and have potential as drugs for limiting corona-virus infections. To search for novel small-molecule frameshift inhibitors with anticoronaviral activity, we computationally screened over 14 million compounds for binding to the SARS-CoV-2 pseudoknot, followed by experimental validation of the top hits for inhibition of -1 PRF and viral replication. We identified multiple potent -1 PRF inhibitors, effective at nM concentrations, some of which significantly suppressed SARS-CoV-2 replication in cell culture. Several compounds also inhibited -1 PRF in multiple representative bat coronaviruses, indicating broad-spectrum activity. These results showcase the promise of viral RNA structures like frameshift-stimulatory pseudoknots as targets for broad-spectrum antiviral drugs.

biophysics↗

bioRxiv2022

Low normal factor V enhances thrombin generation in hemophilia A through a substrate competition mechanism with factor Xa

Bleeding patterns in people with hemophilia A cannot be predicted solely by factor VIII (FVIII) levels. Some of the variance in bleeding may be attributed to differences in plasma protein composition, and specifically other coagulation factors where the normal ranges span 50-150% of the population mean. We recently used a mathematical model of thrombus formation that identified factor V (FV) levels as a strong modifier of thrombin generation in FVIII deficiencies. Counterintuitively, the model predicted low normal FV levels enhanced thrombin generation. Here, we tested this prediction and investigated its mechanism. Thrombin generation in plasma from people with FVIII deficiencies (<5%) were negatively correlated with FV levels. A substrate competition mechanism wherein FV and FVIII compete for activation by FXa during the initiation of coagulation was tested in three models: In a purified system containing only FV, FVIII, and FXa, reducing FV enhanced FVIII activation. In synthetic plasma containing the essential proteins of the extrinsic coagulation pathway, low normal FV levels resulted in enhanced thrombin generation both in the presence or absence of TFPI. In mixture studies using FVIII-deficient human plasma immunodepleted of FV, thrombin generation was enhanced at lower levels of FV. In all models the trend was nonlinear as the effect size was significant at low, but not high, FV levels. Our data show that low normal plasma levels of FV enhance thrombin generation in hemophilia A by reducing FXa substrate competition for FVIII activation and implicate FV levels as a strong modifier of bleeding in hemophilia A. Key PointsO_LILow normal levels of FV enhance thrombin generation in hemophilia A by reducing substate competition for FVIII activation. C_LIO_LIPlasma FV levels are a strong modifier of bleeding in hemophilia A. C_LI

biochemistry↗